Trypanosoma brucei brucei causes the bovine disease Nagana but is non-infectious to humans because of its susceptibility to the cytolytic activity of normal human serum. This activity is due to human haptoglobin related protein (HPR). HPR is found in human sera either as an HDL associated apolipoprotein, trypanosome Lytic factor-i (TLF-l), or as a large protein complex termed TLF-2. The cellular pathway of TLF-l mediated lysis of T b. brucei includes receptor mediated binding to the trypanosome surface, endocytosis and lysosomal targeting. TLF-l kills T b. brucei by destabilization of the lysosomal membrane in an iron dependent reaction that leads to lysosome disruption and cell lysis. The human sleeping sickness parasite Trypanosoma brucei rhodesiense is resistant to TLF-mediated lysis. The mechanism of T b. rhodesiense resistance to TLF-l is associated with a reduction in TLF-l uptake and intracellular trafficking. Expression of a single protein, Serum Resistance Associated (SRA) is sufficient to confer resistance to TLF-l in vitro and in vivo. This protein is a 59 kDa membrane protein that is a member of the VSG gene family. SRA expression is restricted to human sleeping sickness trypanosomes and its expression is transcriptionally regulated. In the proposed studies we will continue to investigate the biochemical and molecular mechanism of J-IPR killing of trypanosomes and the mechanism of SRA protection from TLF-l killing in T b. rhodesiense. The specific aims of this proposal are the following: (1) Determine the structural and biochemical requirements for HPR killing of trypanosomes; (2) Develop a transgenic mouse model for human HPR; (3) Determine the cellular and biochemical mechanisms of SRA inhibition of HPR killing of T b. brucei. Our long-term goals remain to explore the possibilities that modification of HPR or its uptake byhuman sleeping sickness trypanosomes may lead to the identification of novel approaches for treatment of this increasingly important human disease based on this potent innate killing factor.